Page 1235 - Equine Clinical Medicine, Surgery and Reproduction, 2nd Edition

1210 CHAPTER 12

VetBooks.ir 10% buffered neutral formal saline (10× volume of horses, preferring the use of histopathology. Biopsy
samples should be placed in 10% formal saline for
fixative to specimen).
The skin must not be stretched during biopsy.
immunofluorescence testing.
A scalpel or circular biopsy punch is used to sample routine histopathology and in Michel’s medium for
the skin, and fine-toothed forceps or a needle may
be used to carefully remove the biopsy. Ideal speci- Immunohistochemistry staining
men sizes of 6–8 mm or greater will optimise the Immunofluorescent staining was the first method
likelihood of disease pattern recognition. Large developed for immunohistochemistry (IHC) stain-
specimens need to be handled carefully to prevent ing of tissue samples. IHC combines anatomical,
curling and distortion of the specimen in the fixa- immunological and biochemical techniques to image
tive. The pathologist and/or dermatologist should be discrete components in tissues by using appropri-
consulted before taking samples to ensure the best ately labelled antibodies to bind specifically to their
sample sites are selected and the samples fixed prop- target antigens in situ. The advantage of IHC is that
erly for transport. it allows visualisation and documentation of the
The following types of biopsy can be taken: high-resolution distribution and location of spe-
cific cellular components within cells, and within
• A shave biopsy is a thin slice of epidermis their proper histological context by direct, indirect
and small portion of dermis. Suturing is not and double-staining methods. The benefit of more
required. recent IHC methods over immunofluorescence is
• Punch biopsy. A disposable sterile circular that they can often now be accomplished on submit-
6–8 mm punch is most commonly used for ted formalin-fixed tissue samples. This no longer
collection of epidermis, dermis and subcutaneous necessitates stocking of Michel’s medium, which has
fat. If the panniculus muscle or deeper structures a short expiry date.
are a component of the pathology (e.g. lymph
vessels in chronic progressive lymphoedema), the Polymerase chain reaction analysis
sample should be procured by means of a double PCR is a process in which DNA/RNA is extracted
punch technique, using an 8 mm skin punch from a sample, including formalin-fixed biopsies,
biopsy to remove the epidermis, dermis and some and amplified to increase the likelihood of detection
subcutaneous fat, followed by a 6 mm skin punch of viral, rickettsial, bacterial, parasitic, protozoal or
biopsy through the 8 mm opening to acquire fungal organisms. It is also used for detection of cel-
deeper tissue samples including muscle. Suturing lular clonality in patients with suspected neoplasia,
is sometimes required. known as PCR for antigen receptor rearrangement
• A wedge biopsy is a full-thickness section cut (PARR) testing. Real-time PCR is considered the
through the abnormal tissue and includes a small most sensitive and rapid molecular diagnostic assay
piece of normal skin on one edge of the wedge currently in use. Results are typically available in 1–3
for comparison and to orientate the lesion for days. Note, however, that the presence of DNA does
the pathologist. Suturing may be necessary. not reflect the organism’s pathogenicity or the stage
• An excisional biopsy is used when small lesions of disease, it simply indicates its presence within the
are excised whole, with an elliptical incision patient’s sample. Care should be taken to always cor-
using a scalpel, and including all tissue down to relate the PCR findings with clinical disease.
the panniculus muscle. One or more sutures are
placed to close the skin. Antinuclear antibody testing
The ANA titre and LE cell preparation assist in
Immunofluorescence the diagnosis of SLE, which is a rare multisystemic
Tests using these techniques are available in spe- autoimmune disease. The ANA titre detects a com-
cialised pathology laboratories. Some pathologists ponent of the cell nucleus, whereas LE preparation
question the value of direct immunofluorescence in detects antibody to nucleoprotein. While neither test